Stem cells have the unique ability to self-renew and to differentiate into at least one cell type. This unique feature allows stem cells to persist in te adult organism for extended time while continuously producing new differentiating cells. Understanding how stem cells are maintained and differentiated has many promising implications such as stem cell therapies and tissue regenerations. The main objective of this application is to further attempt to understand how the balance between stem cell maintenance and differentiation is controlled. Interference with the pathways that control this balance leads t loss of stem cells if maintenance is defective, and to tumor formation of undifferentiated cells if differentiation is defective. This proposal aims at a systematic, large-scale analysis of stem cell biology that has previously not been possible and promises to identify novel factors that are affecting stem cell maintenance and differentiation. The Drosophila melanogaster female ovary has been widely used as a model system to study stem cells, because of the ability to visualize and follow single stem cells and the many powerful genetic tools available in the field, such as tissue specific knockdowns. I have performed an RNA interference (RNAi) screen to knockdown individual genes in the germline and study the effects on germline stem cell (GSC) maintenance and differentiation. The results of the screen have yield many potential candidates involved with GSCs development. The three specific aims proposed in this application will further characterize the potential candidates affecting GSCs. Aim one will determine the specificity of my RNAi lines that showed promising phenotypes for a particular gene. Aim two will determine what kinds of cells, un/differentiated, are being affected by my candidate gene knockdowns and select genes for further, detailed analysis. Finally, aim three will focus on further characterizing genes and placing them in a GSC developmental pathway. A more complete understanding of the genetic circuits that control stem cells self-renewal and differentiation will be important in particular given of their potential use as therapeutic agents.